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  • The American Society for Biochemistry and Molecular Biology (ASBMB)  (5)
  • The American Physiological Society (APS)  (3)
  • Wiley-Blackwell  (2)
  • 2010-2014  (10)
  • 1
    Publication Date: 2012-12-08
    Description: The microtubule-associated protein targeting protein for Xenopus kinesin-like protein 2 (TPX2) plays a key role in spindle assembly and is required for mitosis in human cells. In interphase, TPX2 is actively imported into the nucleus to prevent its premature activity in microtubule organization. To date, no function has been assigned to nuclear TPX2. We now report that TPX2 plays a role in the cellular response to DNA double strand breaks induced by ionizing radiation. Loss of TPX2 leads to inordinately strong and transient accumulation of ionizing radiation-dependent Ser-139-phosphorylated Histone 2AX (γ-H2AX) at G0 and G1 phases of the cell cycle. This is accompanied by the formation of increased numbers of high intensity γ-H2AX ionizing radiation-induced foci. Conversely, cells overexpressing TPX2 have reduced levels of γ-H2AX after ionizing radiation. Consistent with a role for TPX2 in the DNA damage response, we found that the protein accumulates at DNA double strand breaks and associates with the mediator of DNA damage checkpoint 1 (MDC1) and the ataxia telangiectasia mutated (ATM) kinase, both key regulators of γ-H2AX amplification. Pharmacologic inhibition or depletion of ATM or MDC1, but not of DNA-dependent protein kinase (DNA-PK), antagonizes the γ-H2AX phenotype caused by TPX2 depletion. Importantly, the regulation of γ-H2AX signals by TPX2 is not associated with apoptosis or the mitotic functions of TPX2. In sum, our study identifies a novel and the first nuclear function for TPX2 in the cellular responses to DNA damage.
    Print ISSN: 0021-9258
    Electronic ISSN: 1083-351X
    Topics: Biology , Chemistry and Pharmacology
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  • 2
    Publication Date: 2014-03-14
    Description: Fossil fuel combustion has increased atmospheric CO 2 by ≈ 115 µmol mol -1 since 1750, and decreased its carbon isotope composition (δ 13 C) by 1.7-2 ‰ (the 13 C Suess effect). Because carbon is stored in the terrestrial biosphere for decades and longer, the δ 13 C of CO 2 released by terrestrial ecosystems is expected to differ from the δ 13 C of CO 2 assimilated by land plants during photosynthesis. This isotopic difference between land-atmosphere respiration (δ R ) and photosynthetic assimilation (δ A ) fluxes gives rise to the 13 C land disequilibrium (D). Contemporary understanding suggests that over annual and longer time scales, D is determined primarily by the Suess effect, and thus D is generally positive (δ R  〉 δ A ). A seven-year record of biosphere-atmosphere carbon exchange was used to evaluate the seasonality of δ A and δ R , and the 13 C land disequilibrium, in a subalpine conifer forest. A novel isotopic mixing model was employed to determine the δ 13 C of net land-atmosphere exchange during day and night, and combined with tower-based flux observations to assess δ A and δ R . The disequilibrium varied seasonally, and when flux-weighted was opposite in sign than expected from the Suess effect (D = -0.75 ± 0.21 ‰ or -0.88 ± 0.10 ‰ depending on method). Seasonality in D appeared to be driven by photosynthetic discrimination (Δ canopy ) responding to environmental factors. Possible explanations for negative D include: 1) changes in Δ canopy over decades as CO 2 and temperature have risen, and/or 2) post-photosynthetic fractionation processes leading to sequestration of isotopically-enriched carbon in long-lived pools like wood and soil.
    Print ISSN: 0886-6236
    Electronic ISSN: 1944-9224
    Topics: Biology , Chemistry and Pharmacology , Geography , Geosciences , Physics
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  • 3
    Publication Date: 2014-05-16
    Description: The role of NADPH oxidase (Nox) in both the promotion and impairment of compensatory collateral growth remains controversial because the specific Nox and reactive oxygen species involved are unclear. The aim of this study was to identify the primary Nox and reactive oxygen species associated with early stage compensatory collateral growth in young, healthy animals. Ligation of the feed arteries that form primary collateral pathways in rat mesentery and mouse hindlimb was used to assess the role of Nox during collateral growth. Changes in mesenteric collateral artery Nox mRNA expression determined by real-time PCR at 1, 3, and 7 days relative to same-animal control arteries suggested a role for Nox subunits Nox2 and p47 phox . Administration of apocynin or Nox2ds-tat suppressed collateral growth in both rat and mouse models, suggesting the Nox2/p47 phox interaction was involved. Functional significance of p47 phox expression was assessed by evaluation of collateral growth in rats administered p47 phox small interfering RNA and in p47 phox–/– mice. Diameter measurements of collateral mesenteric and gracilis arteries at 7 and 14 days, respectively, indicated no significant collateral growth compared with control rats or C57BL/6 mice. Chronic polyethylene glycol-conjugated catalase administration significantly suppressed collateral development in rats and mice, implying a requirement for H 2 O 2 . Taken together, these results suggest that Nox2, modulated at least in part by p47 phox , mediates early stage compensatory collateral development via a process dependent upon peroxide generation. These results have important implications for the use of antioxidants and the development of therapies for peripheral arterial disease.
    Print ISSN: 0363-6135
    Electronic ISSN: 1522-1539
    Topics: Medicine
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  • 4
    Publication Date: 2014-11-16
    Description: The enteric nervous system (ENS) is composed of neural crest-derived neurons (also known as ganglion cells) the cell bodies of which are located in the submucosal and myenteric plexuses of the intestinal wall. Intramucosal ganglion cells are known to exist but are rare and often considered ectopic. Also derived from the neural crest are enteric glial cells that populate the ganglia and the associated nerves, as well as the lamina propria of the intestinal mucosa. In Hirschsprung disease (HSCR), ganglion cells are absent from the distal gut because of a failure of neural crest-derived progenitor cells to complete their rostrocaudal migration during embryogenesis. The fate of intramucosal glial cells in human HSCR is essentially unknown. We demonstrate a network of intramucosal cells that exhibit dendritic morphology typical of neurons and glial cells. These dendritic cells are present throughout the human gut and express Tuj1, S100, glial fibrillary acidic protein, CD56, synaptophysin, and calretinin, consistent with mixed or overlapping neuroglial differentiation. The cells are present in aganglionic colon from patients with HSCR, but with an altered immunophenotype. Coexpression of Tuj1 and HNK1 in this cell population supports a neural crest origin. These findings extend and challenge the current understanding of ENS microanatomy and suggest the existence of an intramucosal population of neural crest-derived cells, present in HSCR, with overlapping immunophenotype of neurons and glia. Intramucosal neuroglial cells have not been previously recognized, and their presence in HSCR poses new questions about ENS development and the pathobiology of HSCR that merit further investigation.
    Print ISSN: 0193-1857
    Electronic ISSN: 1522-1547
    Topics: Medicine
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  • 5
    Publication Date: 2014-10-24
    Description: Amateur radio reporting networks, such as the Reverse Beacon Network (RBN), PSKReporter, and the Weak Signal Propagation Network, are powerful tools for remote sensing the ionosphere. These voluntarily constructed and operated networks provide real-time and archival data that could be used for space weather operations, forecasting, and research. The potential exists for the study of both global and localized effects. The capability of one such network to detect space weather disturbances is demonstrated by examining the impacts on RBN-observed HF propagation paths of an X2.9 class solar flare detected by the GOES 15 satellite. Prior to the solar flare, the RBN observed strong HF propagation conditions between multiple continents, primarily Europe, North America, and South America. Immediately following the GOES 15 detection of the solar flare, the number of reported global RBN propagation paths dropped to less than 35% that of prior observations. After the flare, the RBN showed the gradual recovery of HF propagation conditions
    Print ISSN: 1539-4964
    Electronic ISSN: 1542-7390
    Topics: Geosciences , Physics
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  • 6
    Publication Date: 2014-12-02
    Description: Damage to the enteric nervous system (ENS) associated with intestinal inflammation may underlie persistent alterations to gut functions, suggesting that enteric neurons are viable targets for novel therapies. Mesenchymal stem cells (MSCs) offer therapeutic benefits for attenuation of neurodegenerative diseases by homing to areas of inflammation and exhibiting neuroprotective, anti-inflammatory, and immunomodulatory properties. In culture, MSCs release soluble bioactive factors promoting neuronal survival and suppressing inflammation suggesting that MSC-conditioned medium (CM) provides essential factors to repair damaged tissues. We investigated whether MSC and CM treatments administered by enema attenuate 2,4,6-trinitrobenzene-sulfonic acid (TNBS)-induced enteric neuropathy and motility dysfunction in the guinea pig colon. Guinea pigs were randomly assigned to experimental groups and received a single application of TNBS (30 mg/kg) followed by 1 x 10 6 human bone marrow-derived MSCs, 300 μl CM, or 300 μl unconditioned medium 3 h later. After 7 days, the effect of these treatments on enteric neurons was assessed by histological, immunohistochemical, and motility analyses. MSC and CM treatments prevented inflammation-associated weight loss and gross morphological damage in the colon; decreased the quantity of immune infiltrate in the colonic wall ( P 〈 0.01) and at the level of the myenteric ganglia ( P 〈 0.001); prevented loss of myenteric neurons ( P 〈 0.05) and damage to nerve processes, changes in ChAT, and nNOS immunoreactivity ( P 〈 0.05); and alleviated inflammation-induced colonic dysmotility (contraction speed; P 〈 0.001, contractions/min; P 〈 0.05). These results provide strong evidence that both MSC and CM treatments can effectively prevent damage to the ENS and alleviate gut dysfunction caused by TNBS-induced colitis.
    Print ISSN: 0193-1857
    Electronic ISSN: 1522-1547
    Topics: Medicine
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  • 7
    Publication Date: 2014-03-15
    Description: Despite being a major health problem, respiratory syncytial virus (RSV) infections remain without specific therapy. Identification of novel host cellular responses that play a role in the pathogenesis of RSV infection is needed for therapeutic development. The endoplasmic reticulum (ER) stress response is an evolutionarily conserved cellular signaling cascade that has been implicated in multiple biological phenomena, including the pathogenesis of some viral infections. In this study, we investigate the role of the ER stress response in RSV infection using an in vitro A549 cell culture model. We found that RSV infection induces a non-canonical ER stress response with preferential activation of the inositol-requiring enzyme 1 (IRE1) and activated transcription factor 6 (ATF6) pathways with no concomitant significant activation of the protein kinase R-like ER kinase (PERK) pathway. Furthermore, we discovered that IRE1 has an inhibitory effect on RSV replication. Our data characterize, for the first time, the nature of the ER stress response in the setting of RSV infection and identify the IRE1 stress pathway as a novel cellular anti-RSV defense mechanism.
    Print ISSN: 0021-9258
    Electronic ISSN: 1083-351X
    Topics: Biology , Chemistry and Pharmacology
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  • 8
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    The American Society for Biochemistry and Molecular Biology (ASBMB)
    Publication Date: 2012-08-25
    Description: Salmonella enterica serovar typhimurium translocates a glycerophospholipid:cholesterol acyltransferase (SseJ) into the host cytosol after its entry into mammalian cells. SseJ is recruited to the cytoplasmic face of the host cell phagosome membrane where it is activated upon binding the small GTPase, RhoA. SseJ is regulated similarly to cognate eukaryotic effectors, as only the GTP-bound form of RhoA family members stimulates enzymatic activity. Using NMR and biochemistry, this work demonstrates that SseJ competes effectively with Rhotekin, ROCK, and PKN1 in binding to a similar RhoA surface. The RhoA surface that binds SseJ includes the regulatory switch regions that control activation of mammalian effectors. These data were used to create RhoA mutants with altered SseJ binding and activation. This structure-function analysis supports a model in which SseJ activation occurs predominantly through binding to residues within switch region II. We further defined the nature of the interaction between SseJ and RhoA by constructing SseJ mutants in the RhoA binding surface. These data indicate that SseJ binding to RhoA is required for recruitment of SseJ to the endosomal network and for full Salmonella virulence for inbred susceptible mice, indicating that regulation of SseJ by small GTPases is an important virulence strategy of this bacterial pathogen. The dependence of a bacterial effector on regulation by a mammalian GTPase defines further how intimately host pathogen interactions have coevolved through similar and divergent evolutionary strategies.
    Print ISSN: 0021-9258
    Electronic ISSN: 1083-351X
    Topics: Biology , Chemistry and Pharmacology
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  • 9
    Publication Date: 2014-11-29
    Description: Alzheimer disease (AD) is a progressive neurodegenerative disease leading to memory loss. Numerous lines of evidence suggest that amyloid-β (Aβ), a neurotoxic peptide, initiates a cascade that results in synaptic dysfunction, neuronal death, and eventually cognitive deficits. Aβ is generated by the proteolytic processing of the amyloid precursor protein (APP), and alterations to this processing can result in Alzheimer disease. Using in vitro and in vivo models, we identified cyclopamine as a novel regulator of γ-secretase-mediated cleavage of APP. We demonstrate that cyclopamine decreases Aβ generation by altering APP retrograde trafficking. Specifically, cyclopamine treatment reduced APP-C-terminal fragment (CTF) delivery to the trans-Golgi network where γ-secretase cleavage occurs. Instead, cyclopamine redirects APP-CTFs to the lysosome. These data demonstrate that cyclopamine treatment decreases γ-secretase-mediated cleavage of APP. In addition, cyclopamine treatment decreases the rate of APP-CTF degradation. Together, our data demonstrate that cyclopamine alters APP processing and Aβ generation by inducing changes in APP subcellular trafficking and APP-CTF degradation.
    Print ISSN: 0021-9258
    Electronic ISSN: 1083-351X
    Topics: Biology , Chemistry and Pharmacology
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  • 10
    Publication Date: 2014-08-30
    Description: PALB2 links BRCA1 and BRCA2 in homologous recombinational repair of DNA double strand breaks (DSBs). Mono-allelic mutations in PALB2 increase the risk of breast, pancreatic, and other cancers, and biallelic mutations cause Fanconi anemia (FA). Like Brca1 and Brca2, systemic knock-out of Palb2 in mice results in embryonic lethality. In this study, we generated a hypomorphic Palb2 allele expressing a mutant PALB2 protein unable to bind BRCA1. Consistent with an FA-like phenotype, cells from the mutant mice showed hypersensitivity and chromosomal breakage when treated with mitomycin C, a DNA interstrand crosslinker. Moreover, mutant males showed reduced fertility due to impaired meiosis and increased apoptosis in germ cells. Interestingly, mutant meiocytes showed a significant defect in sex chromosome synapsis, which likely contributed to the germ cell loss and fertility defect. Our results underscore the in vivo importance of the PALB2-BRCA1 complex formation in DSB repair and male meiosis.
    Print ISSN: 0021-9258
    Electronic ISSN: 1083-351X
    Topics: Biology , Chemistry and Pharmacology
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